Mechanism of action, potency and efficacy: considerations for cell therapies

One of the most challenging aspects of developing advanced cell therapy products (CTPs) is defining the mechanism of action (MOA), potency and efficacy of the product. This perspective examines these concepts and presents helpful ways to think about them through the lens of metrology. A logical framework for thinking about MOA, potency and efficacy is presented that is consistent with the existing regulatory guidelines, but also accommodates what has been learned from the 27 US FDA-approved CTPs. Available information regarding MOA, potency and efficacy for the 27 FDA-approved CTPs is reviewed to provide background and perspective. Potency process and efficacy process charts are introduced to clarify and illustrate the relationships between six key concepts: MOA, potency, potency test, efficacy, efficacy endpoint and efficacy endpoint test. Careful consideration of the meaning of these terms makes it easier to discuss the challenges of correlating potency test results with clinical outcomes and to understand how the relationships between the concepts can be misunderstood during development and clinical trials. Examples of how a product can be “potent but not efficacious” or “not potent but efficacious” are presented. Two example applications of the framework compare how MOA is assessed in cell cultures, animal models and human clinical trials and reveals the challenge of establishing MOA in humans. Lastly, important considerations for the development of potency tests for a CTP are discussed. These perspectives can help product developers set appropriate expectations for understanding a product’s MOA and potency, avoid unrealistic assumptions and improve communication among team members during the development of CTPs. Supplementary Information The online version contains supplementary material available at 10.1186/s12967-024-05179-7.

Product: Yescarta (axicabtagene ciloleucel) Year approved: 2017 Sponsor: Kite Pharma Description: Genetically modified, antigen-specific autologous T-cells reprogrammed to target cells that express CD19 Indication: For the treatment of adult patients with relapsed or refractory large B-cell lymphoma after two or more lines of systemic therapy, including diffuse large B-cell lymphoma (DLBCL) not otherwise specified, primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, and DLBCL arising from follicular lymphoma Clinical Trial Structure: Single arm (compared to historical data for standard of care) Efficacy Endpoints: Complete remission (as defined complete disappearance of all detectable clinical evidence of disease and symptoms; typically assessed by absence of masses by positron emission tomography (PET), computed tomography (CT) to assess absence of liver/spleen enlargement, bone marrow biopsy) MOA (package insert): "YESCARTA, a CD19-directed genetically modified autologous T-cell immunotherapy, binds to CD19-expressing cancer cells and normal B cells.Studies demonstrated that following anti-CD19 CAR T-cell engagement with CD19-expressing target cells, the CD28 and CD3-zeta co-stimulatory domains activate downstream signaling cascades that lead to T-cell activation, proliferation, acquisition of effector functions and secretion of inflammatory cytokines and chemokines.This sequence of events leads to killing of CD19expressing cells." Potency Testing: • FDA SBRA: i) Cell viability; ii) anti-CD19 CAR expression; iii) redacted • Papadouli et al., 2020: Interferon-γ production by product upon stimulation with CD19+ cells Product: Tecartus (brexucabtagene autoleucel) Year approved: 2020 Sponsor: Kite Pharma Description: TECARTUS is comprised of genetically modified, antigen-specific autologous T-cells reprogrammed to target cells that express CD19, an antigen expressed on the surface of healthy and malignant B cells.The TECARTUS chimeric antigen receptor (CAR) protein has a murine single chain variable fragment (scFv) specific to human CD19 linked to two signaling domains derived from human CD28 and CD3ζ.The CAR protein plays a critical role in TECARTUS function, mediating T-cell activation and anti-tumor effector function following binding of the scFv to CD19.The CAR expressed in TECARTUS is identical to that in YESCARTA (axicabtagene ciloleucel), a CD19-directed genetically modified autologous T-cell immunotherapy approved in 2017 for relapsed or refractory large B cell lymphoma.TECARTUS differs from YESCARTA in that T-cells are enriched during the TECARTUS manufacturing process; T-cell enrichment is not performed during YESCARTA manufacture.Efficacy Endpoint: Score improvements on the Evaluator Wrinkle Assessment Scale and the Subject Wrinkle Assessment Scale

MOA:
• Package Insert: "The mechanism by which LAVIV improves the appearance of nasolabial fold wrinkles is unknown." • FDA SBRA: "The mechanism of action of azficel-T has not been demonstrated.However, each lot is tested to determine that the product consists of viable fibroblasts that produce collagen.The potency of azficel-T is determined by the combination of cell count, viability, identity as fibroblasts and collagen content.The rationale for the choice of these characteristics is based on the premise that fibroblast survival and collagen biosynthesis following injection of azficel-T are likely to be important factors for the improvement in appearance of nasolabial fold wrinkles." • Smith et al, 2012: "The exact mechanism of action of injected autologous fibroblasts remains unknown.They may exert their effect through one of several mechanisms.These could include the direct secretion of increased amounts of collagen and elastin, the induced proliferation of native fibroblasts, the secretion of cofactors that otherwise augment the dermal milieu, or simply multiplication of the transplanted fibroblasts.Most likely is it a combination of several of these processes."Efficacy Endpoints: Ability to regenerate ≥2 mm of keratinized gingiva at 6 months MOA:

Potency
• Package Insert: • "Gintuit does not function as a tissue graft.The mechanism of action by which Gintuit increases keratinized tissue at the treated site has not been identified." • "The active ingredients of GINTUIT are the allogeneic keratinocytes, allogeneic dermal fibroblasts, and bovine Type I collagen.In vitro studies have shown that GINTUIT secretes human growth factors and cytokines, and contains extracellular matrix proteins.Growth factors, cytokines, and extracellular matrix proteins are known to be involved in wound repair and regeneration." • FDA Briefing Document, BLA 125400, November 17, 2011: "The Committee noted that the histology assay is a good measure of the structural integrity of the product, however, the assay is not an adequate, sensitive measure of biological activity.While the exact biological metric that is most appropriate for product potency remains unclear, the Committee discussed that it would be appropriate to include cytokine assays given the current understanding of product function."Potency Testing:

Potency
• FDA SBRA: i) Cell number; ii) redacted; iii) redacted Description: Allogeneic pancreatic islet cellular therapy.LANTIDRA is the first marketed cell-based therapy made from deceased allogeneic donor pancreatic islets of Langerhans (cluster of cells within the pancreas) for the treatment of T1D in adults who are unable to approach target HbA1c because of current repeated episodes of severe hypoglycemia despite intensive diabetes management and education.LANTIDRA is composed of mixed populations of endocrine cells, including beta cells that produce insulin.At least 30% of the product is made of insulin-producing beta cells.Together, the cells composing LANTIDRA regulate blood glucose levels through secretion of hormones in response to glucose stimulation.LANTIDRA is a suspension of islet cells administered through the hepatic portal vein.

Indication:
The treatment of adults with Type 1 diabetes who are unable to approach target HbA1c because of current repeated episodes of severe hypoglycemia despite intensive diabetes management and education.Description (Package Insert): AMTAGVI (lifileucel) is a tumor-derived autologous T cell immunotherapy comprised of a suspension of tumor-derived T cells for intravenous infusion.AMTAGVI is manufactured from resected patient tumor tissue prosected from one or more tumor lesions.Immune cells derived from a patient's tumor(s) are expanded in cell culture, washed, formulated as a cell suspension, and cryopreserved.The product must pass a sterility test before release for shipping as a frozen suspension in 1 to 4 patient-specific infusion bag(s) in individual protective metal cassettes.The product is thawed prior to administration back into the same patient.AMTAGVI is composed primarily of T cells of the CD4+T and CD8+T cell lineages.AMTAGVI may also contain monocytes and other lymphocytes, including B cells and NK cells.AMTAGVI may contain viable melanoma tumor cells from the original tumor tissue used to manufacture the product.
Indication: Indicated for the treatment of adult patients with unresectable or metastatic melanoma previously treated with a PD-1 blocking antibody, and if BRAF V600 mutation positive, a BRAF inhibitor with or without a MEK inhibitor.

Clinical Trial Structure: Single arm
Efficacy Endpoint: Objective response rate according to the RECIST guidelines (Response Evaluation Criteria In Solid Tumors), which is the fraction of patients demonstrating a complete response or partial response.Response is based on tumor load, which is assessed by measuring the longest axis of lesions identified during a baseline examination.The size of lesions can be measured by various means depending on the location: physical examination, chest x-ray, computed tomography, magnetic resonance imaging, ultrasound, etc.The RECIST criteria are as follows:  Complete response-the disappearance of all target lesions;  Partial response-at least a 30% decrease in the sum of the longest diameter of target lesions  Progressive disease-at least a 20% increase in the sum of the longest diameter of target lesions  Stable disease-neither sufficient shrinkage to qualify for partial response nor sufficient increase to qualify for progressive disease MOA (Package Insert): "The specific mechanism of action of AMTAGVI (lifileucel) is unknown." Potency Testing (SBRA): i) redacted, ii) redacted, iii) redacted, iv) redacted, v) dose (total viable cells), vi) redacted and vii) redacted.

:
Relapsed/refractory mantle cell lymphoma Clinical Trial Structure: Single arm (compared to historical data for standard of care) Efficacy Endpoint: Objective response rate [typically assessed by positron emission tomography-computed tomography (PET-CT) for presence of lesions, bone marrow biopsy, lumbar puncture for examination of cerebral spinal fluid (CSF)] MOA (package insert): "TECARTUS, a CD19-directed genetically modified autologous T-cell immunotherapy, binds to CD19-expressing cancer cells and normal B cells.Studies demonstrated that following anti-CD19 CAR T-cell engagement with CD19-expressing target cells, the CD28 and CD3-zeta co-stimulatory domains activate downstream signaling cascades that lead to T-cell activation, proliferation, acquisition of effector functions, and secretion of inflammatory cytokines and chemokines.This sequence of events leads to killing of CD19expressing cells."Potency Testing (FDA SBRA): i) Cell viability; ii) anti-CD19 CAR expression; iii) redacted Testing (FDA SBRA): i) Cell count; ii) cell viability; iii) collagen production by the cells Allogeneic cultured keratinocytes and fibroblasts in bovine collagen Indication: For topical application to a surgically created vascular wound bed in the treatment of adult mucogingival conditions (not intended to provide root coverage) Clinical Trial Structure: Two arm [Gintuit compared to standard of care in the same patient (gingival graft taken from the subject's palate)] Testing (FDA SBRA): Histology (hematoxylin and eosin staining) with morphological assessments: epidermal coverage, epidermal development, basal cell layer keratinocyte viability, suprabasal cell layer keratinocyte viability, dermal thickness, fibroblast density, and matrix integrity Product: MACI (matrix-induced autologous chondrocyte implantation) Year approved: 2016 Sponsor: Vericel Description: Autologous cultured chondrocytes on porcine collagen membrane Indication: Full-thickness cartilage defects of the knee Clinical Trial Structure: Two arm [MACI compared to standard of care (microfracture)] Efficacy Endpoints: Knee injury and Osteoarthritis Outcome Score (KOOS) at 2 yrs MOA (Package Insert): "No clinical pharmacology studies have been conducted with MACI and a mechanism of action has not been established." Unrelated donor hematopoietic progenitor cell transplantation procedures in conjunction with an appropriate preparative regimen for hematopoietic and immunologic reconstitution in patients with disorders affecting the hematopoietic system that are inherited, acquired, or result from myeloablative treatment."Hematopoietic stem/progenitor cells from HPC, Cord Blood migrate to the bone marrow where they divide and mature.The mature cells are released into the bloodstream, where some circulate and others migrate to tissue sites, partially or fully restoring blood counts and function, including immune function, of blood-borne cells of marrow origin."Unrelated donor hematopoietic progenitor cell transplantation procedures in conjunction with an appropriate preparative regimen for hematopoietic and immunologic reconstitution in patients with disorders affecting the hematopoietic system that are inherited, acquired, or result from myeloablative treatment."Hematopoietic stem/progenitor cells from HPC, Cord Blood migrate to the bone marrow where they divide and mature.The mature cells are released into the bloodstream, where some circulate and others migrate to tissue sites, partially or fully restoring blood counts and function, including immune function, of blood-borne cells of marrow origin."Unrelated donor hematopoietic progenitor cell transplantation procedures in conjunction with an appropriate preparative regimen for hematopoietic and immunologic reconstitution in patients with disorders affecting the hematopoietic system that are inherited, acquired, or result from myeloablative treatment."Hematopoietic stem/progenitor cells from HPC, Cord Blood migrate to the bone marrow where they divide and mature.The mature cells are released into the bloodstream, where some circulate and others migrate to tissue sites, partially or fully restoring blood counts and function, including immune function, of blood-borne cells of marrow origin."Unrelated donor hematopoietic progenitor cell transplantation procedures in conjunction with an appropriate preparative regimen for hematopoietic and immunologic reconstitution in patients with disorders affecting the hematopoietic system that are inherited, acquired, or result from myeloablative treatment."Hematopoietic stem/progenitor cells from HPC, Cord Blood migrate to the bone marrow where they divide and mature.The mature cells are released into the bloodstream, where some circulate and others migrate to tissue sites, partially or fully restoring blood counts and function, including immune function, of blood-borne cells of marrow origin."Allogeneic cord blood hematopoietic progenitor cell therapy Indication: Unrelated donor hematopoietic progenitor cell transplantation procedures in conjunction with an appropriate preparative regimen for hematopoietic and immunologic reconstitution in patients with disorders affecting the hematopoietic system that are inherited, acquired, or result from myeloablative treatment."Hematopoietic stem/progenitor cells from HPC, Cord Blood migrate to the bone marrow where they divide and mature.The mature cells are released into the bloodstream, where some circulate and others migrate to tissue sites, partially or fully restoring blood counts and function, including immune function, of blood-borne cells of marrow origin."Allogeneic cord blood hematopoietic progenitor cell therapy Indication: Unrelated donor hematopoietic progenitor cell transplantation procedures in conjunction with an appropriate preparative regimen for hematopoietic and immunologic reconstitution in patients with disorders affecting the hematopoietic system that are inherited, acquired, or result from myeloablative treatment."Hematopoietic stem/progenitor cells from HPC, Cord Blood, migrate to the bone marrow where they divide and mature.The mature cells are released into the bloodstream, where some circulate and others migrate to tissue sites, partially or fully restoring blood counts and function, including immune function, of blood-borne cells of marrow origin."Unrelated donor hematopoietic progenitor cell transplantation procedures in conjunction with an appropriate preparative regimen for hematopoietic and immunologic reconstitution in patients with disorders affecting the hematopoietic system that are inherited, acquired, or result from myeloablative treatment."Hematopoietic stem/progenitor cells from HPC, Cord Blood migrate to the bone marrow where they divide and mature.The mature cells are released into the bloodstream, where some circulate and others migrate to tissue sites, partially or fully restoring blood counts and function, including immune function, of blood-borne cells of marrow origin."Unrelated donor hematopoietic progenitor cell transplantation procedures in conjunction with an appropriate preparative regimen for hematopoietic and immunologic reconstitution in patients with disorders affecting the hematopoietic system that are inherited, acquired, or result from myeloablative treatment."Hematopoietic stem/progenitor cells from HPC, Cord Blood migrate to the bone marrow where they divide and mature.The mature cells are released into the bloodstream, where some circulate and others migrate to tissue sites, partially or fully restoring peripheral blood counts and function, including immune function, of blood-borne cells of marrow origin.""While the precise mechanism of action is unknown, PROVENGE is designed to induce an immune response targeted against PAP, an antigen expressed in most prostate cancers.During ex vivo culture with PAP-GM-CSF, APCs take up and process the recombinant target antigen into small peptides that are then displayed on the APC surface." References• Summary Basis for Regulatory Action -Tecartus,FDA, 2020.Accessed April 27, 2023:https://www.fda.gov/vaccines-blood-biologics/cellular-gene-therapy-products/tecartus-brexucabtageneautoleucel•Package Insert -Tecartus, FDA, 2020.Accessed April 27, 2023: https://www.fda.gov/vaccines-bloodbiologics/cellular-gene-therapy-products/tecartus-brexucabtagene-autoleucel• Wang M, Munoz J, Goy A, Locke FL, Jacobson CA, Hill BT, Timmerman JM, Holmes H, Jaglowski S, Flinn IW, McSweeney PA, Miklos DB, Pagel JM, Kersten MJ, Milpied N, Fung H, Topp MS, Houot R, Beitinjaneh A, Peng W, Zheng L, Rossi JM, Jain RK, Rao AV, Reagan PM.KTE-X19 CAR T-Cell Therapy in Relapsed or Refractory Mantle-Cell Lymphoma.N Engl J Med 2020;382(14):1331-1342. https://doi.org/10.1056/NEJMoa1914347• Assessment Report, European Medicines Agency, Committee for Medicinal Products for Human Use, 2020.• FDA SBRA: i) Redacted • EMA Assessment Report: Interferon-γ production by product upon stimulation with BCMA+ cells Product: Carvykti (ciltacabtagene autoleucel) Year approved: 2023 Sponsor: Janssen Biotech Description: B-cell maturation antigen (BCMA)-directed genetically modified autologous T-cell immunotherapy Indication: Adult patients with relapsed or refractory multiple myeloma after four or more prior lines of therapy, including a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody Clinical Trial Structure: Single arm (compared to historical data for standard of care) Efficacy Endpoints: Survival and objective response rate (as defined by negative response for disease evaluations: i) myeloma protein measurements in serum and urine; ii) serum calcium corrected for albumin; iii) bone marrow examination; iv) skeletal survey; v) documentation of extramedullary plasmacytomas) MOA (Package Insert): "CARVYKTI is a BCMA-directed, genetically modified autologous T-cell immunotherapy, which involves reprogramming a patient's own T-cells with a transgene encoding a chimeric antigen receptor (CAR) that identifies and eliminates cells that express BCMA.The CARVYKTI CAR protein features two BCMAtargeting single-domain antibodies designed to confer high avidity against human BCMA, a 4-1BB costimulatory domain and a CD3-zeta (CD3ζ) signaling cytoplasmic domain.Upon binding to BCMA-expressing cells, the CAR promotes T-cell activation, expansion, and elimination of target cells."Potency Testing (FDA SBRA): i) CAR expression from viable T-cells; ii) redacted Product: Hemacord Year approved: 2011 Sponsor: New York Blood Center Description: Allogeneic cord blood hematopoietic progenitor cell therapy Indication: Clinical Trial Structure: Single arm (compared to historical data for standard of care) Efficacy Endpoints: Hematologic reconstitution (neutrophils, platelets, erythrocytes) MOA (Package Insert): Product: HPC Cord Blood -Clinimmune Labs Year approved: 2012 Sponsor: University of Colorado Cord Blood Bank Description: Allogeneic cord blood hematopoietic progenitor cell therapy Indication: Sponsor: Duke University School of Medicine, Carolinas Cord Blood Bank Description: Allogeneic cord blood hematopoietic progenitor cell therapy Indication: References • Summary Basis for Regulatory Action (SBRA), Clevecord, FDA, 2016.Accessed April 27, 2023: https://www.fda.gov/vaccines-blood-biologics/cellular-gene-therapy-products/clevecord-hpc-cord-blood• Package Insert -Clevecord, FDA, 2016.Accessed April 27, 2023: https://www.fda.gov/vaccines-bloodbiologics/cellular-gene-therapy-products/clevecord-hpc-cord-blood• Laughlin MJ, Barker J, Bambach B, Koc ON, Rizzieri DA, Wagner JE, Gerson SL, Lazarus HM, Cairo M, Stevens CE, Rubinstein P, Kurtzberg J. Hematopoietic engraftment and survival in adult recipients of umbilical-cord blood from unrelated donors.N Engl J Med. 2001 Jun 14;344(24):1815-22.https://doi.org/10.1056/NEJM200106143442402• Laughlin MJ, Eapen M, Rubinstein P, Wagner JE, Zhang MJ, Champlin RE, Stevens C, Barker JN, Gale RP, Lazarus HM, Marks DI, van Rood JJ, Scaradavou A, Horowitz MM.Outcomes after transplantation of cord blood or bone marrow from unrelated donors in adults with leukemia.N Engl J Med. 2004 Nov 25;351(22):2265-75.https://doi.org/10.1056/NEJMoa041276• Cornetta K, Laughlin M, Carter S, Wall D, Weinthal J, Delaney C, Wagner J, Sweetman R, McCarthy P, Chao N. Umbilical cord blood transplantation in adults: results of the prospective Cord Blood Transplantation (COBLT).Biol Blood Marrow Transplant 2005;11(2):149-60.https://doi.org/10.1016/j.bbmt.2004.11.020Product: HPC Cord Blood -MD Anderson Year approved: 2018 Sponsor: MD Anderson Cord Blood Bank Description: Allogeneic cord blood hematopoietic progenitor cell therapy Indication: "In vitro studies have shown that Sratagraft secretes human growth factors and cytokines and contains human ECM proteins.Growth factors, cytokines, and ECM are known to be involved in wound repair and regeneration." "Pancreatic islets regulate blood glucose levels through secretion of multiple hormones in response to increases and decreases in blood glucose.Endocrine cells within pancreatic islets release insulin, glucagon, somatostatin, pancreatic peptide, and ghrelin.Insulin stimulates glucose uptake by peripheral tissues; glucagon mobilizes glucose from the liver into circulation; somatostatin inhibits both α-and β-cell secretions; pancreatic peptide inhibits pancreatic exocrine secretion; and ghrelin inhibits insulin secretion.The primary mechanism of action of LANTIDRA is believed to be secretion of insulin by infused(transplanted) βcells.